Verifying Pluripotency of iPSCs

To proof pluripotency, iPSCs (CMGANTi003-A passage 16 and CMGANTi004-A passage 15) were differentiated into the three embryonic germ layers (mesoderm, endoderm and ectoderm) using the StemMACS Trilineage Differentiation Kit (Miltenyi Biotec) according to manufacturer’s protocol at 37 °C, 5 % CO₂, 20 % O₂. On day seven, cells were collected for RNA extraction and cDNA synthesis. Expression of the selected germ layer markers (Table 1) was verified using RT-qPCR as described above (Table 2).

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De Kinderen P., Peeters S., Rabaut L., Mortier G., Ponsaerts P., Loeys B., Verstraeten A, & Meester J.A. (2023). IPSC reprogramming of two patients with spondyloepimetaphyseal dysplasia (SEMD, biglycan type). Stem Cell Research, 67, 103024.

Publication 2023

StemMACS Trilineage Differentiation Kit

Cdna Cells Ectoderm Embryonic Endoderm Germ layer Ipscs Mesoderm Synthesis

Corresponding Organization : University of Antwerp

Other organizations : KU Leuven, Radboud University Nijmegen, Radboud University Medical Center

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Variable analysis

independent variables

Differentiation protocol (using the StemMACS Trilineage Differentiation Kit)

dependent variables

Expression of germ layer markers

control variables

Incubation temperature (37 °C)
CO2 concentration (5%)
Oxygen concentration (20%)

controls

Positive controls: Not explicitly mentioned
Negative controls: Not explicitly mentioned

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